Innate defense against HIV Infection of CD4+ chemokine co-receptor+ targets by HIV is aided and abetted by the proficiency of HIV in eliminating or neutralizing host cell-derived defensive molecules. Among these innate protective molecules, a family of intracellular apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like (APOBEC) cytidine deaminases is constitutively expressed, but inactivated by HIV viral infectivity factor (Vif). Recently, we demonstrated that type I Interferon, a cytokine with a plethora of functions in innate and adaptive immunity and a potent inhibitor of HIV in vitro and in vivo, exerts its anti-viral activity, at least in part by inducing APOBEC3 family members. The ability of IFN to augment cytidine deaminases offered the possibility that the balance between virus and target cell might be altered in favor of the host. Further characterization of transcriptional profiles induced by IFN using microarrays, with the intention to identify and dissociate retroviral counter-maneuvers from associated toxicities, revealed multiple molecules with suspected anti-viral activity, including IL-27. To establish whether IFN toxicity might be sidestepped through the use of downstream IL-27 against HIV, we examined whether IL-27 directly regulated cytidine deaminases. Dissecting the underlying regulatory events uncovered an initial IL-27-dependent induction of IFN, which in turn, induces APOBEC3, inhibited by IFN receptor blockade. Although the anti-HIV effects of IFN have been demonstrated, efficacy and safety in untreated HIV-infected subjects without chronic viral hepatitis was unknown. Therefore, untreated HIV-infected volunteers without viral hepatitis received weekly pegylated IFNalpha2a for 12 weeks (collaborative studies with NIAID). Changes in HIV RNA, CD4 counts, pharmacokinetics, pharmacodynamic measurements of 2,5 oligoadenylate synthetase (OAS) activity, induction of IFN inducible genes (IFIG) and lymphoproliferative responses (LPA) were measured. Patients with larger increases in OAS tended to have greater decreases in viral load, as did those with higher IFIG induction, whereas those with high IFIG levels at baseline had the lowest subsequent IFIG induction. Based on these clinical studies, peg-IFN has significant anti-HIV activity in HIV-mono-infected patients and this anti-HIV effect correlated with OAS protein and IFIG induction. In ongoing studies, we are defining the mechanisms of reduced viral burden by evaluating specific IFIG and their contribution to inhibition of HIV, including APOBEC, BST/tetherin, and TRIM22. Although IFN therapy reduces viral burden in HIV/HCV co-infected, and also as we have now shown, in HIV-mono-infected individuals, this cytokine can also lead to immune dysfunction and toxicities, limiting its utility. Through detailed mapping of IFN receptor binding sites, we have generated IFN hybrids and mutants in collaboration with investigators at NIAID and determined that structural changes in helix C that influence receptor interactions alter its ability to limit retroviral replication. Our data show a differential ability of the IFN constructs to block HIV replication when compared to wild type IFN, and the directional magnitude of HIV inhibition correlated with levels of APOBEC3 gene expression. As a marker of toxicity, we demonstrated that certain of these mutants induced reduced expression of indoleamine 2,3-dioxygenase (IDO) compared to parental IFN. Subsequent to binding with distinct affinities to the common type I IFN receptor complex (IFNAR), the mutants trigger discreet or shared intracellular signaling pathways (Jak/Stat/PI3K/NFkB) leading to antiviral regulation that may be dissociated from underlying toxic effects, such as IDO. By exploring the structure and function of type I interferon relative to its ability to induce APOBEC and other anti-viral molecules, it may become possible to design novel IFN-related molecules, which preserve its beneficial roles in anti-viral and anti-tumor activity while reducing toxicities that arise in the clinical administration of this potent immunomodulator.